This work is aimed at exploring regulatory mechanisms involved in normal differentiation and malignant transformation, using the B16 melanoma model system involving melanotic and amelanotic malignant clones (and amelanotic nonmalignant clones derived from the melanotic clone after continuous growth in the presence of one microgram of 5-bromodeoxyuridine [BrdUrd] per ml). The BrdUrd clone C3471 immunizes syngeneic mice against the parental tumor. The leukocyte subpopulations elicited by immunization with BrdUrd clone C3471 include macrophages, T cells, and possibly natural killer cells, which cooperate in rejection of the tumorigenic parental melanoma clone (B559). We have isolated and cloned T lymphocytes from normal mice immunologically stimu-lated with BrdUrd clone C3471. These T cells have grown in culture for more than one year with-out interleukin-2 (IL-2) and do not produce IL-2 either constitutively or after stimulation, nor do they produce interferon. Parental and cloned T cells form lymphomas when inoculated into immunosuppressed mice. The T cells and tumors derived from them produce C-type ecotropic retrovirus which readily infects and grows productively in virus-free mouse fibroblasts and melanoma cells. Cocultivation of infected cells with uninfected fibroblasts, melanoma, or XC cells results in the formation of syncytia within 3 to 6 hrs in the presence of Amphotericin B; without Amphotericin B, syncytia form efficiently only in XC cells. The virus-producing lymphocytes are capable of producing a host response in inoculated mice, which can lead to regression of lymphomas and can cause rejection of inoculated melanoma cells. We have also found that we can efficiently transfer selectable and unselectable genes into differentiated and undifferentiated mouse melanoma clones using calcium phosphate coprecipitation. Frequencies of transferent colony formation with melanoma recipients approach or surpass mouse LTK- cells after treatment with either of two codominantly selectable plasmid vectors, pSV2gpt or pGCcos3neo. We applied this system successfully to DNA-mediated transfer of a gene specifying a 130,000 molecular weight glycoprotein (gp 130) from human melanoma cells to one of our mouse melanoma clones at a frequency of 3x10-4. By continuing these approaches we hope to increase our understanding of regulatory mechanisms in differentiation, cell-virus interactions in stimulating neoplastic transformation, and cellular interactions in immune surveillance. (M)